scholarly journals Functional screening of a human saliva metagenomic DNA reveal novel resistance genes against sodium hypochlorite and chlorhexidine

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Johannes Wigand ◽  
Supathep Tansirichaiya ◽  
Endre Winje ◽  
Mohammed Al-Haroni
Keyword(s):  
Sodium Hypochlorite ◽  
Resistance Genes ◽  
Antimicrobial Agents ◽  
Metagenomic Library ◽  
Bioinformatic Analysis ◽  
Oral Microbiome ◽  
Oral Microbiota ◽  
Functional Screening ◽  
Metagenomic Dna ◽  
Dental Practices

Abstract Objective Many sections of the health care system are facing a major challenge making infectious disease problematic to treat; antimicrobial resistance (AMR). Identification and surveillance of the resistome have been highlighted as one of the strategies to overcome the problem. This study aimed to screen for AMR genes in an oral microbiota, a complex microbial system continuously exposed to antimicrobial agents commonly used in dental practice. Materials and methods As a significant part of the oral microbiome cannot be conventionally cultured, a functional metagenomic approach was chosen. The human oral metagenomic DNA was extracted from saliva samples collected from 50 healthy volunteers in Norway. The oral metagenomic library was then constructed by ligating partially digested oral metagenome into pSMART BAC vector and introducing into Escherichia coli. The library was screened against antimicrobials in dental practices. All resistant clones were selected and analyzed. Results Screening of the oral metagenomic library against different antimicrobials detected multiple clones with resistance against chlorhexidine, triclosan, erythromycin, tetracycline, and sodium hypochlorite. Bioinformatic analysis revealed both already known resistance genes, including msr, mef(A), tetAB(46), and fabK, and genes that were not previously described to confer resistance, including recA and accB conferring resistance to sodium hypochlorite and chlorhexidine, respectively. Conclusion Multiple clones conferring resistance to antimicrobials commonly used in dental practices were detected, containing known and novel resistant genes by functional-based metagenomics. There is a need for more studies to increase our knowledge in the field.

scholarly journals Identification of unknown acid-resistant genes of oral microbiotas in patients with dental caries using metagenomics analysis

AMB Express ◽  
2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Xi Cheng ◽  
Fuming He ◽  
Ping Sun ◽  
Qianming Chen
Keyword(s):  
Dental Caries ◽  
High Throughput Sequencing ◽  
Resistance Mechanism ◽  
Metagenomic Library ◽  
Acid Resistance ◽  
Bioinformatic Analysis ◽  
Oral Microbiota ◽  
E Coli ◽  
Resistant Genes ◽  
Oral Microbes

AbstractAcid resistance is critical for the survival of bacteria in the dental caries oral micro-environment. However, there are few acid-resistant genes of microbiomes obtained through traditional molecular biology experimental techniques. This study aims to try macrogenomics technologies to efficiently identify acid-resistant genes in oral microbes of patients with dental caries. Total DNA was extracted from oral microbiota obtained from thirty dental caries patients and subjected to high-throughput sequencing. This data was used to build a metagenomic library, which was compared to the sequences of two Streptococcus mutant known acid-resistant genes, danK and uvrA, using a BLAST search. A total of 19 and 35 unknown gene sequences showed similarities with S. mutans uvrA and dnaK in the metagenomic library, respectively. Two unknown genes, mo-dnaK and mo-uvrA, were selected for primer design and bioinformatic analysis based on their sequences. Bioinformatics analysis predicted them encoding of a human heat-shock protein (HSP) 70 and an ATP-dependent DNA repair enzyme, respectively, closely related with the acid resistance mechanism. After cloning, these genes were transferred into competent Escherichia coli for acid resistance experiments. E. coli transformed with both genes demonstrated acid resistance, while the survival rate of E. coli transformed with mo-uvrA was significantly higher in an acidic environment (pH = 3). Through this experiment we found that identify unknown acid-resistant genes in oral microbes of patients with caries by establishing a metagenomic library is very efficient. Our results provide an insight into the mechanisms and pathogenesis of dental caries for their treatment without affecting oral probiotics.

scholarly journals Oral Microbiota Identifies Patients in Early Onset Rheumatoid Arthritis

2021 ◽  
Vol 9 (8) ◽  
pp. 1657
Author(s):  
Anders Esberg ◽  
Linda Johansson ◽  
Ingegerd Johansson ◽  
Solbritt Rantapää Dahlqvist
Keyword(s):  
Rheumatoid Arthritis ◽  
Early Onset ◽  
Amplicon Sequencing ◽  
Oral Bacteria ◽  
Oral Microbiome ◽  
Rrna Gene ◽  
Oral Microbiota ◽  
Disease Manifestation ◽  
Periodontal Status ◽  
Quality Register

Rheumatoid arthritis (RA) is the most common autoimmune inflammatory disease, and single periodontitis-associated bacteria have been suggested in disease manifestation. Here, the oral microbiota was characterized in relation to the early onset of RA (eRA) taking periodontal status into consideration. 16S rRNA gene amplicon sequencing of saliva bacterial DNA from 61 eRA patients without disease-modifying anti-rheumatic drugs and 59 matched controls was performed. Taxonomic classification at 98.5% was conducted against the Human Oral Microbiome Database, microbiota functions were predicted using PICRUSt, and periodontal status linked from the Swedish quality register for clinically assessed caries and periodontitis. The participants were classified into three distinct microbiota-based cluster groups with cluster allocation differences by eRA status. Independently of periodontal status, eRA patients had enriched levels of Prevotella pleuritidis, Treponema denticola, Porphyromonas endodontalis and Filifactor alocis species and in the Porphyromonas and Fusobacterium genera and functions linked to ornithine metabolism, glucosylceramidase, beta-lactamase resistance, biphenyl degradation, fatty acid metabolism and 17-beta-estradiol-17-dehydrogenase metabolism. The results support a deviating oral microbiota composition already in eRA patients compared with healthy controls and highlight a panel of oral bacteria that may be useful in eRA risk assessment in both periodontally healthy and diseased persons.

scholarly journals Pre and Probiotics Involved in the Modulation of Oral Bacterial Species: New Therapeutic Leads in Mental Disorders?

2021 ◽  
Vol 9 (7) ◽  
pp. 1450
Author(s):  
Yoann Maitre ◽  
Rachid Mahalli ◽  
Pierre Micheneau ◽  
Alexis Delpierre ◽  
Marie Guerin ◽  
...  
Keyword(s):  
Systematic Review ◽  
Central Nervous System ◽  
Nervous System ◽  
Oral Health ◽  
Mental Disorders ◽  
Meta Analysis ◽  
Bacterial Species ◽  
Oral Microbiome ◽  
Oral Microbiota ◽  
Therapeutic Leads

This systematic review aims to identify probiotics and prebiotics for modulating oral bacterial species associated with mental disorders. Using the Preferred Reporting Items for Systematic Reviews and Meta-Analysis guideline, we search the electronic MEDLINE database published till January 2021 to identify the studies on probiotics and/or prebiotics for preventing and treating major oral dysbiosis that provokes mental disorders. The outcome of the search produces 374 records. After excluding non-relevant studies, 38 papers were included in the present review. While many studies suggest the potential effects of the oral microbiota on the biochemical signalling events between the oral microbiota and central nervous system, our review highlights the limited development concerning the use of prebiotics and/or probiotics in modulating oral dysbiosis potentially involved in the development of mental disorders. However, the collected studies confirm prebiotics and/or probiotics interest for a global or targeted modulation of the oral microbiome in preventing or treating mental disorders. These outcomes also offer exciting prospects for improving the oral health of people with mental disorders in the future.

scholarly journals Oral microbial dysbiosis and its performance in predicting oral cancer

2020 ◽  
Author(s):  
Shih-Chi Su ◽  
Lun-Ching Chang ◽  
Hsien-Da Huang ◽  
Chih-Yu Peng ◽  
Chun-Yi Chuang ◽  
...  
Keyword(s):  
Oral Cancer ◽  
Fusobacterium Nucleatum ◽  
Risky Behaviors ◽  
Health Condition ◽  
Oral Microbiome ◽  
Oral Microbiota ◽  
Microbial Dysbiosis ◽  
Anti Cancer ◽  
Betel Quid Chewing ◽  
Male Patients

Abstract Dysbiosis of oral microbiome may dictate the progression of oral squamous cell carcinoma (OSCC). Yet, the composition of oral microbiome fluctuates by saliva and distinct sites of oral cavity and is affected by risky behaviors (smoking, drinking and betel quid chewing) and individuals’ oral health condition. To characterize the disturbances in the oral microbial population mainly due to oral tumorigenicity, we profiled the bacteria within the surface of OSCC lesion and its contralateral normal tissue from discovery (n = 74) and validation (n = 42) cohorts of male patients with cancers of the buccal mucosa. Significant alterations in the bacterial diversity and relative abundance of specific oral microbiota (most profoundly, an enrichment for genus Fusobacterium and the loss of genus Streptococcus in the tumor sites) were identified. Functional prediction of oral microbiome shown that microbial genes related to the metabolism of terpenoids and polyketides were differentially enriched between the control and tumor groups, indicating a functional role of oral microbiome in formulating a tumor microenvironment via attenuated biosynthesis of secondary metabolites with anti-cancer effects. Furthermore, the vast majority of microbial signatures detected in the discovery cohort was generalized well to the independent validation cohort, and the clinical validity of these OSCC-associated microbes was observed and successfully replicated. Overall, our analyses reveal signatures (a profusion of Fusobacterium nucleatum CTI-2 and a decrease in Streptococcus pneumoniae) and functions (decreased production of tumor-suppressive metabolites) of oral microbiota related to oral cancer.

scholarly journals Functional Genomic Identification of Cadmium Resistance Genes from a High GC Clone Library by Coupling the Sanger and PacBio Sequencing Strategies

Genes ◽  
2019 ◽  
Vol 11 (1) ◽  
pp. 7
Author(s):  
Jinghao Chen ◽  
Chao Xing ◽  
Xin Zheng ◽  
Xiaofang Li
Keyword(s):  
High Throughput ◽  
Resistance Genes ◽  
Sanger Sequencing ◽  
Genomic Library ◽  
Full Length ◽  
Host Cells ◽  
Full Genome Sequence ◽  
Functional Screening ◽  
Functional Genomic ◽  
Pacbio Sequencing

Functional (meta) genomics allows the high-throughput identification of functional genes in a premise-free way. However, it is still difficult to perform Sanger sequencing for high GC DNA templates, which hinders the functional genomic exploration of a high GC genomic library. Here, we developed a procedure to resolve this problem by coupling the Sanger and PacBio sequencing strategies. Identification of cadmium (Cd) resistance genes from a small-insert high GC genomic library was performed to test the procedure. The library was generated from a high GC (75.35%) bacterial genome. Nineteen clones that conferred Cd resistance to Escherichia coli subject to Sanger sequencing directly. The positive clones were in parallel subject to in vivo amplification in host cells, from which recombinant plasmids were extracted and linearized by selected restriction endonucleases. PacBio sequencing was performed to obtain the full-length sequences. As the identities, partial sequences from Sanger sequencing were aligned to the full-length sequences from PacBio sequencing, which led to the identification of seven unique full-length sequences. The unique sequences were further aligned to the full genome sequence of the source strain. Functional screening showed that the identified positive clones were all able to improve Cd resistance of the host cells. The functional genomic procedure developed here couples the Sanger and PacBio sequencing methods and overcomes the difficulties in PCR approaches for high GC DNA. The procedure can be a promising option for the high-throughput sequencing of functional genomic libraries, and realize a cost-effective and time-efficient identification of the positive clones, particularly for high GC genetic materials.

scholarly journals Identification and Characterization of a Halotolerant, Cold-Active Marine Endo-β-1,4-Glucanase by Using Functional Metagenomics of Seaweed-Associated Microbiota

2014 ◽  
Vol 80 (16) ◽  
pp. 4958-4967 ◽  
Author(s):  
Marjolaine Martin ◽  
Sophie Biver ◽  
Sébastien Steels ◽  
Tristan Barbeyron ◽  
Murielle Jam ◽  
...  
Keyword(s):  
Metagenomic Library ◽  
Functional Screening ◽  
Sample Collection ◽  
Prolonged Incubation ◽  
Content Type ◽  
Cold Active ◽  
Esterase Loci ◽  
Collection Period ◽  
Identification And Characterization

ABSTRACTA metagenomic library was constructed from microorganisms associated with the brown algaAscophyllum nodosum. Functional screening of this library revealed 13 novel putative esterase loci and two glycoside hydrolase loci. Sequence and gene cluster analysis showed the wide diversity of the identified enzymes and gave an idea of the microbial populations present during the sample collection period. Lastly, an endo-β-1,4-glucanase having less than 50% identity to sequences of known cellulases was purified and partially characterized, showing activity at low temperature and after prolonged incubation in concentrated salt solutions.

scholarly journals Antimicrobial Susceptibilities of Oral Isolates of Abiotrophia and Granulicatella According to the Consensus Guidelines for Fastidious Bacteria

Medicines ◽  
2018 ◽  
Vol 5 (4) ◽  
pp. 129 ◽  
Author(s):  
Taisei Kanamoto ◽  
Shigemi Terakubo ◽  
Hideki Nakashima
Keyword(s):  
Dna Hybridization ◽  
Vitamin B6 ◽  
Antimicrobial Agents ◽  
Oral Microbiota ◽  
Phenotypic Characteristics ◽  
Granulicatella Adiacens ◽  
Susceptibility Profiles ◽  
Susceptibility Rate ◽  
Type Strains ◽  
Culture Negative

Background: The genera Abiotrophia and Granulicatella, previously known as nutritionally variant streptococci (NVS), are fastidious bacteria requiring vitamin B6 analogs for growth. They are members of human normal oral microbiota, and are supposed to be one of the important pathogens for so-called “culture-negative” endocarditis. Methods: The type strains and oral isolates identified, by using both phenotypic profiles and the DNA–DNA hybridization method, were examined for susceptibilities to 15 antimicrobial agents including penicillin (benzylpenicillin, ampicillin, amoxicillin, and piperacillin), cephem (cefazolin, ceftazidime, ceftriaxone, and cefaclor), carbapenem (imipenem), aminoglycoside (gentamicin), macrolide (erythromycin), quinolone (ciprofloxacin), tetracycline (minocycline), glycopeptide (vancomycin), and trimethoprim-sulfamethoxazole complex. The minimum inhibitory concentration and susceptibility criterion were determined, according to the consensus guideline from the Clinical and Laboratory Standards Institute. Results: Isolates of Abiotrophia defectiva were susceptible to ampicillin, amoxicillin ceftriaxone, cefaclor, imipenem, ciprofloxacin, and vancomycin. Isolates of Granulicatella adiacens were mostly susceptible to benzylpenicillin, ampicillin, amoxicillin, cefazolin, ceftriaxone, imipenem, minocycline, and vancomycin. The susceptibility profile of Granulicatella elegans was similar to that of G. adiacens, and the susceptibility rate was higher than that of G. adiacens. Conclusions: Although Abiotrophia and Granulicatella strains are hardly distinguishable by their phenotypic characteristics, their susceptibility profiles to the antimicrobial agents were different among the species. Species-related differences in susceptibility of antibiotics should be considered in the clinical treatment for NVS related infections.

scholarly journals Functional screening for resistance genes against trichothecenes in the library of Saccharomyces cerevisiae deletion mutants

2013 ◽  
Vol 63 (1) ◽  
pp. 9-15 ◽  
Author(s):  
Naoko TAKAHASHI-ANDO ◽  
Akira TANAKA ◽  
Yohsuke SEKIMOTO ◽  
Kohta YAMAUCHI ◽  
Akinobu ECHIGO ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Resistance Genes ◽  
Deletion Mutants ◽  
Functional Screening ◽  
Screening For Resistance

scholarly journals Inducible Macrolide Resistance inCorynebacterium jeikeium

2001 ◽  
Vol 45 (7) ◽  
pp. 1982-1989 ◽  
Author(s):  
Adriana E. Rosato ◽  
Bonnie S. Lee ◽  
Kevin A. Nash
Keyword(s):  
Drug Resistance ◽  
Resistance Genes ◽  
Broad Spectrum ◽  
Antimicrobial Agents ◽  
Opportunistic Pathogen ◽  
Macrolide Resistance ◽  
The Other ◽  
Broad Spectrum Resistance ◽  
Neutropenic Patients ◽  
Group 3

ABSTRACT Corynebacterium jeikeium is an opportunistic pathogen primarily of immunocompromised (neutropenic) patients. Broad-spectrum resistance to antimicrobial agents is a common feature of C. jeikeium clinical isolates. We studied the profiles of susceptibility of 20 clinical strains of C. jeikeium to a range of antimicrobial agents. The strains were separated into two groups depending on the susceptibility to erythromycin (ERY), with one group (17 strains) representing resistant organisms (MIC > 128 μg/ml) and the second group (3 strains) representing susceptible organisms (MIC ≤ 0.25 μg/ml). The ERY resistance crossed to other members of the macrolide-lincosamide-streptogramin B (MLSb) group. Furthermore, this resistance was inducible with MLSb agents but not non-MLSb agents. Expression of ERY resistance was linked to the presence of an allele of the class X erm genes,erm(X)cj, with >93% identity to other ermgenes of this class. Our evidence indicates that erm(X)cj is integrated within the chromosome, which contrasts with previous reports for the plasmid-associated erm(X) genes found inC. diphtheriae and C. xerosis. In 40% ofC. jeikeium strains, erm(X)cj is present within the transposon, Tn5432. However, in the remaining strains, the components of Tn5432 (i.e., the erm and transposase genes) have separated within the chromosome. The rearrangement of Tn5432 leads to the possibility that the other drug resistance genes have become included in a new composite transposon bound by the IS1249 elements.

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